GENOME MAINTENANCE BY THE MOUSE HUS1 CHECKPOINT GENE Cancers arise due to the sequential accumulation of mutations in growth regulatory genes. The safeguarding of genomic integrity therefore plays a critical role in tumor suppression. Genome maintenance is achieved in part through cell cycle checkpoints, which prevent the accumulation of genome damage by regulating cell cycle progression, apoptosis, and DNA repair. In mammals, the proteins Atr, Hus1, and Chk1 comprise a checkpoint pathway that responds to specific DNA lesions as well as DNA replication arrest. The long-term goal of the research described in this proposal is to determine how this Hus1-dependent checkpoint pathway maintains genomic stability and contributes to normal development and tumor suppression. In aim one of this proposal, we will identify requirements for Husl in DNA damage signaling and cell cycle checkpoints in primary mouse cells. In aim two, we will establish whether Husl inactivation in adult mice promotes tumorigenesis and impairs tissue-specific DNA damage responses. These objectives will be accomplished using a new system for the regulated inactivation of Hus1 that bypasses the requirement for this essential pathway during murine embryonic development. While complete inactivation of the Hus1-dependent checkpoint mechanism leads to severe genomic instability, additional evidence suggests that a partial deficiency in this pathway can cause developmental defects and promote tumor formation. In aim three, we will utilize a novel hypomorphic Hus1 allele that expresses greatly reduced levels of Hus1 to test whether partial impairment of the Hus 1-dependent checkpoint pathway causes genomic instability and altered genotoxic stress responses in cultured cells and mice. Together, the proposed studies will reveal how a key mammalian checkpoint pathway operates throughout the cell cycle and additionally will define the roles of this checkpoint mechanism in tumor suppression and in vivo genome maintenance.